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Determination of different gonadotropin synthesis mechanisms in trophoblast and choriocarcinoma cells

滋养层细胞和绒毛膜癌细胞中不同促性腺激素合成机制的测定

基本信息

批准号：
63570133
负责人：
SAKAKIBARA Ryuzo
金额：
$ 1.34万
依托单位：
Faculty of Pharmaceutical Sciences, Nagasaki University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1988
资助国家：
日本
起止时间：
1988 至 1989
项目状态：
已结题

项目摘要

Human chorionic gonadotropin (hCG) is a glycoprotein hormone consisting of noncovalently bonded subunits, alpha and beta. Both subunits contain two Asn(N)-linked sugar chains, in addition, the beta-subunit contains four Ser(O)-linked sugar chains. It has been suggested that altered glycosylation is induced in both alpha- and beta-subunits of hCG produced by choriocarcinoma. The objective of this study was to determine the nature of the structural differences of hormones synthesized in the cells and secreted as well as the uncombined subunits in normal trophoblasts and a choriocarcinoma cell line (BeWo).The results obtained from this study are as follows:1. Intracellular forms of hCG subunits in normal trophoblast cells were analyzed by immunobinding method. It was revealed that hCG subunits accumulate in trophoblast cells as predominant intermediates (immature forms) containing high-mannose sugar chains. On the other hand, Ser-linked sugar chain was not found in the immature beta-subunit.2. The immature hCG subunits were purified, separately, and their physicochemical properties were determined. N-terminal amino acid sequence of each immature subunit was identical to respective mature form. The purified immature subunits were recombined each other and reconstituted an immature hCG. The apparent molecular weights of the immature hCG found in trophoblast cells as well as hCG reconstituted with the purified immature subunits were larger than those expected. It was suggested that the immature hCG is present in trophoblast cells as an oligomer.3. The structural characteristics of hCG and its subunits in choriocarcinoma, BeWo cells, were compared to those of normal trophoblast cells. It was found that both protein and sugar chain structures of hCG subunits of BeWo cells were differed from those of trophoblast cells.

人绒毛膜促性腺激素(HCG)是一种糖蛋白激素，由非共价键的α和β亚基组成。这两个亚基都含有两个Asn(N)连接的糖链，另外，β-亚基还包含四个Ser(O)连接的糖链。已有研究表明，绒毛膜癌产生的hCG的α和β亚基均发生了糖基化改变。本研究旨在探讨正常滋养层细胞和绒毛膜癌细胞系(BeWo)细胞内合成和分泌的激素及其未结合亚基的结构差异。结果如下：1.用免疫结合方法分析正常滋养层细胞内hCG亚基的形态。研究表明，hCG亚基在滋养层细胞中以含有高甘露糖链的中间产物(未成熟形式)的形式积累。另一方面，在未成熟的β-亚基中没有发现丝氨酸连接的糖链。对未成熟的hCG亚基分别进行纯化，并测定其理化性质。各未成熟亚基N端氨基酸序列与成熟亚基一致。纯化的未成熟亚基相互重组，重组为未成熟的hCG。在滋养层细胞中发现的未成熟hCG以及由纯化的未成熟亚基重组的hCG的表观分子量比预期的要大。提示未成熟的hCG以寡聚体形式存在于滋养层细胞中。比较了绒癌BeWo细胞和正常滋养层细胞hCG及其亚基的结构特征。结果表明，BeWo细胞hCG亚基的蛋白质和糖链结构与滋养层细胞不同。